using comments_template() in custom single-portfolio.php

When registering a custom post type you need to enable comments. In the arguments defining the CPT, you need to include something like this:

'supports' => array( 'title', 'editor', 'author', 'thumbnail', 'excerpt', 'comments' )

The example on the Codex is shows where this option needs to go, this page also shows a list of other options you may or may not want to include: http://codex.wordpress.org/Function_Reference/register_post_type#Example